Team:UT-Tokyo/CTCD/Content/Modeling

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Revision as of 00:08, 18 October 2014

Characterization of BBa_K747096

During DNA assembly, we found that this promoter was activated in Escherichia coli. Then we decided to characterize this promoter activity by comparing with constitutive promoter (BBa_J23101) using GFP (BBa_E0040) (Figure 1).
As seen in Figure 1, this promoter shows weaker activity than BBa_J23101.

<legend style="width:400px;float:left;margin-left:50px;margin-right:10px;">Fig. 1The activity of BBa_K747096GFP was activated by 501nm excitation laser.
a)Measured when OD600 is 0.9~1.1.
b)Measured after cultured O/N.
In order to calculate RPU (relative promoter unit [1]), we performed real-time measurement of GFP fluorescence (Figure 2). We measured activity of GFP and OD600, which are necessary for calculation of RPU.</legend> <legend style="width:400px;float:left;margin-left:0px;margin-right:0px;">Fig. 2Real-time measurement of BBa_K747096
a)Activity of GFP (activated by 488nm excitation laser).
b)OD600.
We calculated RPU(Relative Promoter Unit) of BBa_K747096 from the Fig. 2 using the equation (1).</legend> <img src="" class="math" height="60px" style="clear:both;"/>

This equation can be used only when Fluorescence per cell is in steady state. In this experiment, this condition was fulfilled as shown in Table1.

<tbody>

</tbody>

Table 1 : Fluorescence / OD
	K747096	J23101
t=4h	92.62	13.14
t=6h	99.37	19.86

From Table1, we can get RPU of BBa_K747096:

RPU = 5.8

[1]:https://2010.igem.org/Team:Kyoto/LearnMore#Relative_Promoter_Unit_.28RPU.29

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 					<img src = "https://static.igem.org/mediawiki/2014/f/f0/Graph%28assay5%29_maxwiki.png" class = "figure" style="width:400px; float:left;padding-left:0px;" />
 					<p style="clear:both"></p>
-					<legend style="width:400px;float:left;margin-left:50px;margin-right:60px;"><b>Fig. 1</b>The activity of BBa_K747096GFP was activated by 501nm excitation laser. <br>a)Measured when OD600 is 0.9~1.1.<br>b)Measured after cultured O/N.<br />In order to calculate RPU (relative promoter unit [1]), we performed real-time measurement of GFP fluorescence (Figure 2). We measured activity of GFP and OD600, which are necessary for calculation of RPU.</legend>
+					<legend style="width:400px;float:left;margin-left:50px;margin-right:10px;"><b>Fig. 1</b>The activity of BBa_K747096GFP was activated by 501nm excitation laser. <br>a)Measured when OD600 is 0.9~1.1.<br>b)Measured after cultured O/N.<br />In order to calculate RPU (relative promoter unit [1]), we performed real-time measurement of GFP fluorescence (Figure 2). We measured activity of GFP and OD600, which are necessary for calculation of RPU.</legend>
-					<legend style="width:400px;float:left;"><b>Fig. 2</b>Real-time measurement of BBa_K747096<br>a)Activity of GFP (activated by 488nm excitation laser).<br>b)OD600. <br />We calculated RPU(Relative Promoter Unit) of BBa_K747096 from the Fig. 2 using the equation (1).</legend>
+					<legend style="width:400px;float:left;margin-left:0px;margin-right:0px;"><b>Fig. 2</b>Real-time measurement of BBa_K747096<br>a)Activity of GFP (activated by 488nm excitation laser).<br>b)OD600. <br />We calculated RPU(Relative Promoter Unit) of BBa_K747096 from the Fig. 2 using the equation (1).</legend>
 					<img src="https://static.igem.org/mediawiki/2014/d/d1/Cocoa_equation_1.png" class="math" height="60px" style="clear:both;"/>
 					<p>This equation can be used only when Fluorescence per cell is in steady state. In this experiment, this condition was fulfilled as shown in Table1.</p>